Integrins are a group of cell surface glycoproteins which mediate cell adhesion and therefore are useful mediators of cell adhesion interactions which occur during various biological processes. Integrins are heterodimers composed of noncovalently linked .alpha. and .beta. polypeptide subunits. Currently eleven different .alpha. subunits have been identified and six different .beta. subunits have been identified. The various .alpha. subunits can combine with various .beta. subunits to form distinct integrins.
The integrin identified as .alpha..sub.v.beta..sub.3 (also known as the vitronectin receptor) has been identified as an integrin which plays a role in various conditions or disease states including tumor metastasis, solid tumor growth (neoplasia), osteoporosis, Paget's disease, humoral hypercalcemia of malignancy, angiogenesis, including tumor angiogenesis, retinopathy, including macular degeneration, arthritis, including rheumatoid arthritis, periodontal disease, psoriasis and smooth muscle cell migration (e.g. restenosis). Additionally, it has been found that such agents would be useful as antivirals, antifungals and antimicrobials. Thus, compounds which selectively inhibit or antagonize .alpha..sub.v.beta..sub.3 would be beneficial for treating such conditions.
It has been shown that the .alpha..sub.v.beta..sub.3 integrin and other .alpha..sub.v containing integrins bind to a number of Arg-Gly-Asp (RGD) containing matrix macromolecules. Compounds containing the RGD sequence mimic extracellular matrix ligands so as to bind to cell surface receptors. However, it is also known that RGD peptides in general are non-selective for RGD dependent integrins. For example, most RGD peptides which bind to .alpha..sub.v.beta..sub.3 also bind to .alpha..sub.v.beta..sub.5, .alpha..sub.v.beta..sub.1 and .alpha..sub.IIb.beta..sub.3. Antagonism of platelet .alpha..sub.IIb.beta..sub.3 (also known as the fibrinogen receptor) is known to block platelet aggregation in humans. In order to avoid bleeding side-effects when treating the conditions or disease states associated with the integrin .alpha..sub.v.beta..sub.3, it would be beneficial to develop compounds which are selective antagonists of .alpha..sub.v.beta..sub.3 as opposed to .alpha..sub.IIb.beta..sub.3.
Tumor cell invasion occurs by a three step process: 1) tumor cell attachment to extracellular matrix; 2) proteolytic dissolution of the matrix; and 3) movement of the cells through the dissolved barrier. This process can occur repeatedly and can result in metastases at sites distant from the original tumor.
Seftor et al. (Proc. Natl. Acad. Sci. USA, Vol. 89 (1992) 1557-1561) have shown that the .alpha..sub.v.beta..sub.3 integrin has a biological function in melanoma cell invasion. Montgomery et al., (Proc. Natl. Acad. Sci USA, Vol. 91 (1994) 8856-60) have demonstrated that the integrin .alpha..sub.v.beta..sub.3 expressed on human melanoma cells promotes a survival signal, protecting the cells from apoptosis. Mediation of the tumor cell metastatic pathway by interference with the .alpha..sub.v.beta..sub.3 integrin cell adhesion receptor to impede tumor metastasis would be beneficial.
Brooks et al. (Cell, Vol. 79 (1994) 1157-1164) have demonstrated that antagonists of .alpha..sub.v.beta..sub.3 provide a therapeutic approach for the treatment of neoplasia (inhibition of solid tumor growth) since systemic administration of .alpha..sub.v.beta..sub.3 antagonists causes dramatic regression of various histologically distinct human tumors.
The adhesion receptor integrin .alpha..sub.v.beta..sub.3 was identified as a marker of angiogenic blood vessels in chick and man and therefore such receptor plays a critical role in angiogenesis or neovascularization. Angiogenesis is characterized by the invasion, migration and proliferation of smooth muscle and endothelial cells. Antagonists of .alpha..sub.v.beta..sub.3 inhibit this process by selectively promoting apoptosis of cells in neovasculature. The growth of new blood vessels, or angiogenesis, also contributes to pathological conditions such as diabetic retinopathy and macular degeneration (Adonis et al., Amer. J. Ophthal., Vol. 118, (1994) 445-450) and rheumatoid arthritis (Peacock et al., J. Exp. Med., Vol. 175, (1992), 1135-1138). Therefore, .alpha..sub.v.beta..sub.3 antagonists would be useful therapeutic targets for treating such conditions associated with neovascularization (Brooks et al., Science, Vol. 264, (1994), 569-571).
It has been reported that the cell surface receptor .alpha..sub.v.beta..sub.3 is the major integrin on osteoclasts responsible for attachment to bone. Osteoclasts cause bone resorption and when such bone resorbing activity exceeds bone forming activity it results in osteoporosis (a loss of bone), which leads to an increased number of bone fractures, incapacitation and increased mortality. Antagonists of .alpha..sub.v.beta..sub.3 have been shown to be potent inhibitors of osteoclastic activity both in vitro [Sato et al., J. Cell. Biol., Vol. 111 (1990) 1713-1723] and in vivo [Fisher et al., Endocrinology, Vol. 132 (1993) 1411-1413]. Antagonism of .alpha..sub.v.beta..sub.3 leads to decreased bone resorption and therefore restores a normal balance of bone forming and resorbing activity. Thus it would be beneficial to provide antagonists of osteoclast .alpha..sub.v.beta..sub.3 which are effective inhibitors of bone resorption and therefore are useful in the treatment or prevention of osteoporosis.
The role of the .alpha..sub.v.beta..sub.3 integrin in smooth muscle cell migration also makes it a therapeutic target for prevention or inhibition of neointimal hyperplasia which is a leading cause of restenosis after vascular procedures (Choi et al., J. Vasc. Surg. Vol. 19(1) (1994) 125-34). Prevention or inhibition of neointimal hyperplasia by pharmaceutical agents to prevent or inhibit restenosis would be beneficial.
White (Current Biology, Vol. 3(9)(1993) 596-599) has reported that adenovirus uses .alpha..sub.v.beta..sub.3 for entering host cells. The integrin appears to be required for endocytosis of the virus particle and may be required for penetration of the viral genome into the host cell cytoplasm. Thus compounds which inhibit .alpha..sub.v.beta..sub.3 would find usefulness as antiviral agents.
U.S. Ser. No. 09/034,270 discloses compounds of the following general formula ##STR2##
wherein X and Y are the same or different halo group; R is H or alkyl; and pharmaceutically acceptable salts thereof. Such compounds find use as .alpha..sub.v.beta..sub.3 integrin antagonists.
More specifically, U.S. Ser. No. 09/034,270 discloses the following compounds: ##STR3##
wherein R is H or alkyl; or pharmaceutically acceptable salts thereof. Each of these compounds contains a chiral .beta.-amino acid/ester moiety which is substituted by halogens. In order to prepare such .alpha..sub.v.beta..sub.3 antagonists, it is therefore useful to have methodology to efficiently prepare a chiral .beta.-amino acid/ester.
Chiral chromatography to separate enantiorners is known [Peter et al., Analytica Chimica Acta, 352 (1997) 335-356] and is a possible method for preparing such chiral compounds. The present invention provides a novel method for preparation of chiral .beta. amino esters which are useful in the preparation of .alpha..sub.v.beta. antagonists described above.